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- EMDB-10574: Beta-galactosidase in complex with PETG -

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Basic information

Entry
Database: EMDB / ID: EMD-10574
TitleBeta-galactosidase in complex with PETG
Map dataNone
Sample
  • Complex: Beta-galactosidase
    • Protein or peptide: Beta-galactosidase
  • Ligand: MAGNESIUM ION
  • Ligand: 2-phenylethyl 1-thio-beta-D-galactopyranoside
  • Ligand: water
KeywordsBgal / PETG / SUGAR BINDING PROTEIN
Function / homology
Function and homology information


alkali metal ion binding / lactose catabolic process / beta-galactosidase complex / beta-galactosidase / beta-galactosidase activity / carbohydrate binding / magnesium ion binding / identical protein binding
Similarity search - Function
Glycoside hydrolase, family 2, beta-galactosidase / Beta galactosidase small chain/ domain 5 / Beta-galactosidase, domain 4 / Beta galactosidase small chain / Beta-galactosidase, domain 4 / Beta galactosidase small chain / : / Glycoside hydrolase, family 2, active site / Glycosyl hydrolases family 2 acid/base catalyst. / Glycoside hydrolase, family 2, conserved site ...Glycoside hydrolase, family 2, beta-galactosidase / Beta galactosidase small chain/ domain 5 / Beta-galactosidase, domain 4 / Beta galactosidase small chain / Beta-galactosidase, domain 4 / Beta galactosidase small chain / : / Glycoside hydrolase, family 2, active site / Glycosyl hydrolases family 2 acid/base catalyst. / Glycoside hydrolase, family 2, conserved site / Glycosyl hydrolases family 2 signature 1. / Glycoside hydrolase, family 2 / Glycosyl hydrolases family 2, sugar binding domain / Glycoside hydrolase family 2, catalytic domain / Glycosyl hydrolases family 2, sugar binding domain / Glycosyl hydrolases family 2, TIM barrel domain / Glycoside hydrolase, family 2, immunoglobulin-like beta-sandwich / Glycosyl hydrolases family 2 / Beta-Galactosidase/glucuronidase domain superfamily / Glycoside hydrolase-type carbohydrate-binding / Galactose mutarotase-like domain superfamily / Galactose-binding-like domain superfamily / Glycoside hydrolase superfamily / Immunoglobulin-like fold
Similarity search - Domain/homology
Beta-galactosidase / Beta-galactosidase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.2 Å
AuthorsSaur M / Hartshorn MJ
CitationJournal: Drug Discov Today / Year: 2020
Title: Fragment-based drug discovery using cryo-EM.
Authors: Michael Saur / Michael J Hartshorn / Jing Dong / Judith Reeks / Gabor Bunkoczi / Harren Jhoti / Pamela A Williams /
Abstract: Recent advances in electron cryo-microscopy (cryo-EM) structure determination have pushed the resolutions obtainable by the method into the range widely considered to be of utility for drug discovery. ...Recent advances in electron cryo-microscopy (cryo-EM) structure determination have pushed the resolutions obtainable by the method into the range widely considered to be of utility for drug discovery. Here, we review the use of cryo-EM in fragment-based drug discovery (FBDD) based on in-house method development. We demonstrate not only that cryo-EM can reveal details of the molecular interactions between fragments and a protein, but also that the current reproducibility, quality, and throughput are compatible with FBDD. We exemplify this using the test system β-galactosidase (Bgal) and the oncology target pyruvate kinase 2 (PKM2).
History
DepositionDec 27, 2019-
Header (metadata) releaseJan 15, 2020-
Map releaseJan 15, 2020-
UpdateMay 22, 2024-
Current statusMay 22, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6tte
  • Surface level: 0.05
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10574.png

Downloads & links

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Map

FileDownload / File: emd_10574.map.gz / Format: CCP4 / Size: 35.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNone
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.68 Å/pix.
x 295 pix.
= 200.6 Å		0.68 Å/pix.
x 222 pix.
= 150.96 Å		0.68 Å/pix.
x 143 pix.
= 97.24 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 0.68 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.1434283 - 0.27425525
Average (Standard dev.)0.002424097 (±0.024056187)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions222143295
Spacing143222295
CellA: 97.24 Å / B: 150.96 Å / C: 200.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.680.680.68
M x/y/z143222295
origin x/y/z0.0000.0000.000
length x/y/z97.240150.960200.600
α/β/γ90.00090.00090.000
start NX/NY/NZ1081340
NX/NY/NZ13584349
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS143222295
D min/max/mean-0.1430.2740.002

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Supplemental data

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Additional map: Relion post-process unmasked map

Fileemd_10574_additional.map
AnnotationRelion post-process unmasked map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Relion post-process unmasked map

Fileemd_10574_additional_1.map
AnnotationRelion post-process unmasked map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Relion auto-refine halfmap 1

Fileemd_10574_half_map_1.map
AnnotationRelion auto-refine halfmap 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Relion auto-refine halfmap 2

Fileemd_10574_half_map_2.map
AnnotationRelion auto-refine halfmap 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Beta-galactosidase

EntireName: Beta-galactosidase
Components
  • Complex: Beta-galactosidase
    • Protein or peptide: Beta-galactosidase
  • Ligand: MAGNESIUM ION
  • Ligand: 2-phenylethyl 1-thio-beta-D-galactopyranoside
  • Ligand: water

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Supramolecule #1: Beta-galactosidase

SupramoleculeName: Beta-galactosidase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 464 KDa

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Macromolecule #1: Beta-galactosidase

MacromoleculeName: Beta-galactosidase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: beta-galactosidase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 118.395336 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGSSHHHHHH SSGLVPRGSH MLEDPVVLQR RDWENPGVTQ LNRLAAHPPF ASWRNSEEAR TDRPSQQLRS LNGEWRFAWF PAPEAVPES WLECDLPEAD TVVVPSNWQM HGYDAPIYTN VTYPITVNPP FVPTENPTGC YSLTFNVDES WLQEGQTRII F DGVNSAFH ...String:
MGSSHHHHHH SSGLVPRGSH MLEDPVVLQR RDWENPGVTQ LNRLAAHPPF ASWRNSEEAR TDRPSQQLRS LNGEWRFAWF PAPEAVPES WLECDLPEAD TVVVPSNWQM HGYDAPIYTN VTYPITVNPP FVPTENPTGC YSLTFNVDES WLQEGQTRII F DGVNSAFH LWCNGRWVGY GQDSRLPSEF DLSAFLRAGE NRLAVMVLRW SDGSYLEDQD MWRMSGIFRD VSLLHKPTTQ IS DFHVATR FNDDFSRAVL EAEVQMCGEL RDYLRVTVSL WQGETQVASG TAPFGGEIID ERGGYADRVT LRLNVENPKL WSA EIPNLY RAVVELHTAD GTLIEAEACD VGFREVRIEN GLLLLNGKPL LIRGVNRHEH HPLHGQVMDE QTMVQDILLM KQNN FNAVR CSHYPNHPLW YTLCDRYGLY VVDEANIETH GMVPMNRLTD DPRWLPAMSE RVTRMVQRDR NHPSVIIWSL GNESG HGAN HDALYRWIKS VDPSRPVQYE GGGADTTATD IICPMYARVD EDQPFPAVPK WSIKKWLSLP GETRPLILCE YAHAMG NSL GGFAKYWQAF RQYPRLQGGF VWDWVDQSLI KYDENGNPWS AYGGDFGDTP NDRQFCMNGL VFADRTPHPA LTEAKHQ QQ FFQFRLSGQT IEVTSEYLFR HSDNELLHWM VALDGKPLAS GEVPLDVAPQ GKQLIELPEL PQPESAGQLW LTVRVVQP N ATAWSEAGHI SAWQQWRLAE NLSVTLPAAS HAIPHLTTSE MDFCIELGNK RWQFNRQSGF LSQMWIGDKK QLLTPLRDQ FTRAPLDNDI GVSEATRIDP NAWVERWKAA GHYQAEAALL QCTADTLADA VLITTAHAWQ HQGKTLFISR KTYRIDGSGQ MAITVDVEV ASDTPHPARI GLNCQLAQVA ERVNWLGLGP QENYPDRLTA ACFDRWDLPL SDMYTPYVFP SENGLRCGTR E LNYGPHQW RGDFQFNISR YSQQQLMETS HRHLLHAEEG TWLNIDGFHM GIGGDDSWSP SVSAEFQLSA GRYHYQLVWC QK

UniProtKB: Beta-galactosidase

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Macromolecule #2: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 2 / Number of copies: 4 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #3: 2-phenylethyl 1-thio-beta-D-galactopyranoside

MacromoleculeName: 2-phenylethyl 1-thio-beta-D-galactopyranoside / type: ligand / ID: 3 / Number of copies: 4 / Formula: PTQ
Molecular weightTheoretical: 300.371 Da
Chemical component information

ChemComp-PTQ:
2-phenylethyl 1-thio-beta-D-galactopyranoside

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 1224 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.17 mg/mL
BufferpH: 6.8
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: GRAPHENE OXIDE / Support film - topology: CONTINUOUS
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 562 / Average exposure time: 59.98 sec. / Average electron dose: 65.53 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 136013
Startup modelType of model: EMDB MAP
EMDB ID:

4116

Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.04) / Number images used: 49895
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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